Rapid screening of pyogenic Staphylococcus aureus for confirmation of genus and species, methicillin resistance and virulence factors by using two novel multiplex PCR

Objectives: Emergence of methicillin resistant Staphylococcus aureus [MRSA] is a major medical problem of current era. These bacteria are resistant to most drugs and rapid diagnosis can provide a clear guideline to clinicians. They possess specific virulence factors and relevant information can be very useful. We designed this study to develop multiplex PCRs to provide rapid information

Methods: We studied 60 Staphylococcus aureus isolates and detected methicillin resistance by cefoxitin sensitivity and targeting of mecA gene. After initial studies with uniplex PCRs we optimized two multiplex PCRs with highly reproducible results. The first multiplex PCR was developed to confirm genus, species and methicillin resistance simultaneously, and the second multiplex PCR was for screening of virulence factors

Results: We found 38.33% isolates as methicillin resistant, a-toxin, the major cytotoxic factor, was detected in 40% whereas 6-hemolysin was found in 25% cases. Panton Valentine leucocidin was detected in 8.33% and toxic shock syndrome toxin in5% cases. The results of uniplex and multiplex PCRs were highly compatible

Conclusions: These two multiplex PCRs when run simultaneously can provide vital information about methicillin resistance and virulence status of the isolate within a few hours as compared to several days needed by routine procedures

Bacteriological analysis of drinking water from 100 families of Lahore by membrane filtration technique and chromagar

Water borne diseases are caused by pathogenic microorganisms. In Pakistan, the availability of safe water is only 40% to 60%. Therefore it becomes imperative to determine the bacteriological status of drinking water. A few laboratories perform such an evaluation and, that too, by the old method technique i.e Most Probable Number [MPN]. We evaluated 100 samples of drinking water from some areas of Lahore by the Metnbrane Filtration Technique [MFT] using CHRO-Magar. Using this technique in one step a much large volume of water can be evaluated quantitatively in a short time and with ease. Use of CHROMagar straightaway confirms the presence of Escherichia coli which is accepted universally as the indicator of fecal contamination. It was a cross sectional study. A volume of 100 ml water was filtered under the vacuum pressure through Millipore membrane filters. After filtration, membrane filters were placed on CHRO Magar and incubated at 35°C for 24 hr. Escherichia coli appeared as blue coloured colonies while coliforms yielded colonies of pink colour. Escherichia coli were further identified by API 2OE and confirmed by Eijkman test. Escherichia coli was grown from 42% samples [all Eijkman positive]. Coliform organisms were grown from 54% specimens. It was alarming that 59% of drinking water was unsatisfactory for human consumption